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Mol Med. 2018 Aug 14;24(1):44. doi: 10.1186/s10020-018-0046-1.Loss of Transient Receptor Potential Melastatin 3 ion channel function in natural killer cells from Chronic Fatigue Syndrome/Myalgic Encephalomyelitis patients.Cabanas H1,2, Muraki K3, Eaton N4,5, Balinas C4,5, Staines D4,5, Marshall-Gradisnik S4,5.Author informationAbstractBACKGROUND:
Chronic Fatigue Syndrome (CFS)/ Myalgic Encephalomyelitis (ME) is a debilitating disorder that is accompanied by reduced cytotoxic activity in natural killer (NK) cells. NK cells are an essential innate immune cell, responsible for recognising and inducing apoptosis of tumour and virus infected cells. Calcium is an essential component in mediating this cellular function. Transient Receptor Potential Melastatin 3 (TRPM3) cation channels have an important regulatory role in mediating calcium influx to help maintain cellular homeostasis. Several single nucleotide polymorphisms have been reported in TRPM3 genes from isolated peripheral blood mononuclear cells, NK and B cells in patients with CFS/ME and have been proposed to correlate with illness presentation. Moreover, a significant reduction in both TRPM3 surface expression and intracellular calcium mobilisation in NK cells has been found in CFS/ME patients compared with healthy controls. Despite the functional importance of TRPM3, little is known about the ion channel function in NK cells and the epiphenomenon of CFS/ME. The objective of the present study was to characterise the TRPM3 ion channel function in NK cells from CFS/ME patients in comparison with healthy controls using whole cell patch-clamp techniques.

METHODS:
NK cells were isolated from 12 age- and sex-matched healthy controls and CFS patients. Whole cell electrophysiology recording has been used to assess TRPM3 ion channel activity after modulation with pregnenolone sulfate and ononetin.

RESULTS:
We report a significant reduction in amplitude of TRPM3 current after pregnenolone sulfate stimulation in isolated NK cells from CFS/ME patients compared with healthy controls. In addition, we found pregnenolone sulfate-evoked ionic currents through TRPM3 channels were significantly modulated by ononetin in isolated NK cells from healthy controls compared with CFS/ME patients.

CONCLUSIONS:
TRPM3 activity is impaired in CFS/ME patients suggesting changes in intracellular Ca2+ concentration, which may impact NK cellular functions. This investigation further helps to understand the intracellular-mediated roles in NK cells and confirm the potential role of TRPM3 ion channels in the aetiology and pathomechanism of CFS/ME.

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This picture suggests why the researchers have got so excited about their finding. It shows very big differences between healthy controls in a) and b) compared to patients in c) and d). The test involves shocking cells with PregS and seeing how their ion channels respond.

Here is their conclusion:

We have demonstrated impaired TRPM3 activity in CFS/ME patients through electrophysiological investi- gations in NK cells after modulation with PregS and ononetin. As TRPM3 is widely distributed in the body, particularly brain, eye, cardiovascular system, gastrointestinal system and pancreas, we assert that widespread changes in TRPM3 function in key body systems may contribute to CFS/ME. We suggest changes to Ca2+ ion concentration in the cytosol and intracellular stores result from changes in TRPM3 function, which may impact NK cellular functions. Therefore, Ca2+ signalling pathways could be an alter- native therapeutic target in CFS/ME because of their importance in various cellular processes. This investi- gation confirms the potential role of TRPM3 ion channels in the aetiology and pathomechanism of CFS/ME, and could suggest potential therapeutic tar- gets and/or prognostic markers.

Assuming the finding is real, it is substantial. Could be a fluke, could be researcher bias, could be machine error, or it could be real.

The study has only 6 patients and 6 controls, but of course the p-value of .0001 takes the small sample into account. We don't need to overcorrect for the small sample and reductively dismiss this paper. If you're looking for something obvious, like comparing the size of a watermelon and a walnut, you don't need a huge sample to start to see a strong signal. I will be very interested to hear what people think.

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@Murph I know the preliminary findings of this study were posted before, and I know that I asked about it back then, but now that the entire results are out (or at least I think they are?), can you explain what it means in basic terms? Does it relate to having the Calcium Channel autoantibody or LEMS or is there no relationship whatsoever? I struggle to decipher what these studies are talking about!

:)

@Murph I know the preliminary findings of this study were posted before, and I know that I asked about it back then, but now that the entire results are out (or at least I think they are?), can you explain what it means in basic terms? Does it relate to having the Calcium Channel autoantibody or LEMS or is there no relationship whatsoever? I struggle to decipher what these studies are talking about!

It doesn't say anything either way about auto-antobodies. But I think they may suspect it is genetic. Their previous studies found some relevant gene mutations. I've sent a couple of emails to the researchers asking more questions.

What it says is actually simple because it is very limited: one particular type of channel that gets ions like calcium into and out of our cells is not working properly.

It doesn't really say anything definitive about why, or about the implications, or about treatments. Those are blanks for us to pencil in until future research finds out for sure.

:)

There exists a promoter of TRPM3 callled d -erythro sphingosine. The search function above says this compound has been mentioned on this site only twice.
The word sphingosine got me thinking about the sphingolipids that Naviaux found were all low.

So I took another look at Naviaux's data, from his big metabolomics study. Here's a screengrab of my analysis of that data, with the ratios of different metabolites in patients and controls. A lot of what we're short in is something called ceramides, ( they often have some alphanumeric stuff in brackets after them).

I wanted to remember exactly what those ceramides were, so I googled.

I found this exciting. At least some of the listed ceramides might be types of d -erythro sphingosine. So it is possible our bodies are often low in certain compounds that would stimulate these ion channels to work.

If I'm interpreting this right (a big if - my training is in economics!) there could be a link from Naviaux's work to the Griffith research.

1. This may means the Griffith work might not be as far out on its own branch as we thought.
2. This may mean they're discovering a downstream problem from metabolic disturbance (lack of ceramides mean trpm3 doesn't work)
3. This may mean they're discovering an upstream problem (lack of ceramides is due to exhausation of ceramide supplies in trying to make trpm3 work.)

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Im very excited on any new research around ME/CFS and calicum as i know I have issues with calicum for a couple of reasons.

1/ I have lots of double copy uncommon gene mutations to do with the calicum ion channel, when I looked at my uncommon mutations I was surprised in how much that area stood out, this was even before I knew calicum channels were being connected to ME/CFS. (Im about to see i I can find the print out on them to see if I can see this specific one mentioned in this study among the ones I have mutated).

Hi gingergrrl, yes the raw data from 23andME was used to find out my more uncommon double copy gene mutations. I never though analysed all of it but used a program to pull out the uncommon ones (which someone from this website had made) and then was researching each one to try to find out about the mutations and what they affected. I only got through about a quarter or a third of my uncommon mutations.

and no I have never been tested for calcium channel autoantibodies...
....

I earlier looked through "some" of the pages of I'd analysed of my gene mutation results and in those did not see the one listed i this article but can see I have rs12239772 which is one called CACNA1S which is to do with the calicum channel (associated with hypolcalemic periodic paralysis type 1.. deficiency of calicum in the blood stream... so maybe some of my paralyses bouts during crashes were related to calicum?????) https://en.wikipedia.org/wiki/Cav1.1 "Cav1.1 also known as the calcium channel, voltage-dependent, L type, alpha 1S subunit, (CACNA1S), is a protein which in humans is encoded by the CACNA1S gene.[5]"
"This gene encodes one of the five subunits of the slowly inactivating L-type voltage-dependent calcium channel in skeletal muscle cells."

Anyway I wonder how calicum deals with meditating cells if one has a mutation which interfers with cell binding so maybe even IF it turns out that I havent got this one in this study mutated, maybe I could still get similar problem with calicum processes...

There exists a promoter of TRPM3 callled d -erythro sphingosine. The search function above says this compound has been mentioned on this site only twice.View attachment 29138
The word sphingosine got me thinking about the sphingolipids that Naviaux found were all low.

So I took another look at Naviaux's data, from his big metabolomics study. Here's a screengrab of my analysis of that data, with the ratios of different metabolites in patients and controls. A lot of what we're short in is something called ceramides, ( they often have some alphanumeric stuff in brackets after them).

View attachment 29140
I found this exciting. At least some of the listed ceramides might be types of d -erythro sphingosine. So it is possible our bodies are often low in certain compounds that would stimulate these ion channels to work.

If I'm interpreting this right (a big if - my training is in economics!) there could be a link from Naviaux's work to the Griffith research.

1. This may means the Griffith work might not be as far out on its own branch as we thought.
2. This may mean they're discovering a downstream problem from metabolic disturbance (lack of ceramides mean trpm3 doesn't work)
3. This may mean they're discovering an upstream problem (lack of ceramides is due to exhausation of ceramide supplies in trying to make trpm3 work.)

Senior Member

Senior Member

that's very interesting, I am also very low for pregnenolone(TRPM3 agonist), and I took a lot of it as supplement, but it never made any difference for me.
I am not convinced that the TRPM3 channels are involved in my case, but many other ion channels are involved in intracellular calcium homeostasis, so I think TRPM3 is only a small peace of the picture.

on the contrary, I think that every scientific papers studying our intracellular calcium homeostasis can provide new treatement options for us. I mean, not something to treat the root of our disease, but something to improve our clinical outcome.

Intracellular calcium is dependant on many different ion channels, many hormons, many signals, so it can be disturbed by a bunch of different pathways, which could explain why we can observe subgroups in the ME syndrome.

I have low T3/TSH (both in the limit of the normal ranges though), and I tryed several protocols with T4 or T3, and for now only T3 works, and not more than a small dose once a day.

It works very fast, which is consistent with a direct effect of T3 on ion channels.

I also now experience clearly a kind of paralysis if I take any macrolides antibiotics (erythro, clarythro, azithromycine), so I am now convinced my neuro-muscular junction doesn't work properly.

I earlier looked through "some" of the pages I'd analysed of my gene mutation results and in those did not see the one listed i this article but can see I have rs12239772 which is one called CACNA1S which is to do with the calicum channel (associated with hypolcalemic periodic paralysis type 1.. deficiency of calicum in the blood stream... so maybe some of my paralyses bouts during crashes were related to calicum?????) https://en.wikipedia.org/wiki/Cav1.1 "Cav1.1 also known as the calcium channel, voltage-dependent, L type, alpha 1S subunit, (CACNA1S), is a protein which in humans is encoded by the CACNA1S gene.[5]" "This gene encodes one of the five subunits of the slowly inactivating L-type voltage-dependent calcium channel in skeletal muscle cells."

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Reading this groups last paper (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5217865/) I recall being one of the most difficult paper's to follow and understand. I just tried to refresh my memory of it, and quite honestly I still don't follow parts of the experiments. Particularly why PregS requires thapsigargin as well to increase or restore effector function in NK cells. If they are saying in order to reverse the effects, they ALSO need to inhibit the function of an ion channeel/pump in the ER which has nothing to do with TRPM3, then why is that? Is the suggestion that PregS by itself is not enough stimulation to increase the intracellular gradient enough to restore effector function? I don't know, I might have expected the PregS alone to provide more restoration of effector function.

Anyway, I consider myself to be reasonably well equipped to read most papers, but I honestly cannot understand the nuts and bolts of this properly. It's difficult to be excited by paper's that I don't understand well.

Previous papers identified:

NK effector Function is known to be reduced in ME/CFS

TRPM3 Single Nucleotide deficiencies (SNP's) in TRPM3 genes have previously been identified in ME/CFS patients as a possible common genetic mutation

This group demonstrated:

Intracellular stores of Calcium Ions in ME/CFS patient NK cells are abnormal, leading to reduced effector function in NK cells

Initially I was quite unsure about these findings, because in my head I was thinking "wouldn't CFS manifest as a primary immune disorder if NK cell effector function is reduced?", but I guess that under this model, it's not that NK function is non-existant, it's just that it's reduced. Still, I would have thought this would manifest as serious illness. Particularly bad viral infections, and greatly increased risk of cancers of all types. It's still something that I wonder about, and that is why I'm not sure about these findings.

Well yes, it may be that most of these findings are indeed downstream effects. It has bothered me a great deal as well, but I started to look at things a bit differently. If we take for example type 1 diabetes, the uncontrolled glucose and lack of insulin is a downstream effect and the actual cause is destruction of insulin producing beta cells in the pancreas. But diabetes as of today cannot be treated at this root cause level, which would be to stop destruction of beta cells instead of just injecting insulin. So in order to have an effective treatment, it's not always necessary to treat the root cause, even if that ultimately would produce a better result (I read they are actually trying to find ways to regenerate these beta cells as well).